Sensor Housing and Reagent Chemistry

ABSTRACT

A sensor comprises a sensor housing, having a channel; a porous substrate, in the channel; an analysis chemistry reagent, on the porous substrate; and a nozzle, in fluid connection with the channel. The porous substrate fills a cross section of the channel, and the cross-sectional area of the channel at the porous substrate is greater than the cross-sectional area at the nozzle.

BACKGROUND

Methods which permit real time detection of Pb²⁺ and other metal ionsare very important in the fields of environmental monitoring, clinicaltoxicology, wastewater treatment, and industrial process monitoring andcan lead to preventative measures or lower risks associated with metalcontaminants. However, traditional methods of detecting metal ions arecumbersome, often requiring samples collected in the field to be broughtback and analyzed in a laboratory setting. Methods are needed for realtime detection and monitoring of metal ions in industrial and biologicalsystems.

Fluorescence spectroscopy is a technique well suited for detection ofvery small concentrations of analytes. Fluorescence provides significantsignal amplification, since a single fluorophore can absorb and emitmany photons, leading to strong signals even at very low concentrations.In addition, the fluorescence time-scale is fast enough to allowreal-time monitoring of concentration fluctuations. Fluorescentproperties only respond to changes related to the fluorophore, andtherefore can be highly selective. Also, fluorometers, for measuringfluorescence signals, are commercially available.

SUMMARY

In a first aspect, the present invention is a sensor, comprising asensor housing, having a channel; a porous substrate, in the channel; ananalysis chemistry reagent, on the porous substrate; and a nozzle, influid connection with the channel. The porous substrate fills a crosssection of the channel, and the cross-sectional area of the channel atthe porous substrate is greater than the cross-sectional area at thenozzle.

In a second aspect, the present invention is a kit, comprising a box;and a plurality of sensors, and a plurality of cuvettes. One of theplurality of sensors and one of the plurality of cuvettes are each in afirst sealed bag of a plurality of first sealed bags.

In a third aspect, the present invention is a method of detecting anion, comprising flowing a sample fluid through the channel of a sensor,to produce a product; collecting the product; and detecting the presenceof the product.

The following definitions are included to provide a clear and consistentunderstanding of the specification and claims.

The term “sample” is defined as a composition suspected of containingthe analyte of interest that will be subjected to analysis. Typically, asample for analysis is in liquid form, or can be converted into liquidform, and preferably the sample is an aqueous composition. A sample maybe from any source, such as an industrial sample from a waste stream, ora biological sample such as blood, urine or saliva. Other examplesinclude drinking water, paint, or paint chips. A sample may be treated,such as by extract, dilution or filtration, or it may be a reconstitutedprecipitate from an industrial or biological source.

The term “analyte” is defined as one or more substances potentiallypresent in a sample, for which the analysis tests. An analysis for ananalyte determines the presence, quantity or concentration, of theanalyte in the sample.

The term “analysis chemistry reagents” refers to one or more reagents,that when reacted with a sample containing an analyte, produce avisualization species. Preferably, the visualization species is producedin proportion to the amount or concentration of the analyte. Analysischemistry reagents preferably include a reactor and a substrate. The“reactor” is at least one compound, moiety and/or material; the“substrate” is also at least one compound, moiety and/or material. Whenthe reactor and the substrate are mixed with the analyte, they willreact to produce a visualization species. As used herein, the term“produce” includes forming by chemical reaction, as well as releasingfrom being bound or attached to something else. Preferably, the reactoris specific for an analyte, and the substrate is specific for a reactor.Preferably, the substrate includes a label. The reactor and thesubstrate may be attached, for example covalently or by hydrogen bonding(hybridization).

The term “visualization species” is a compound, moiety or material thatcan be detected, such as a fluorescent compound or a colored compound. Avisualization species includes a label, which is that part of thevisualization species that allows for detection, for example a coloredlabel (such as a dye or a colored particle, including semiconductornanoparticles (quantum dots)), a fluorescent label (such as fluorescentcompound) or a magnetic label (such as a magnetic particle). Preferably,the label of the visualization species originated as the label of thesubstrate. It is possible for the visualization species and thesubstrate to be the same.

The term “specifically bind” means that binding between the two thingsis more favored binding, as compared to most other members of the sameclass or genus. For example, the binding between an antibody specificfor an antigen, and the antigen; and hybridization between twocomplementary strands of DNA; are both examples of specific binding.

The term “aptamer” refers to nucleic acid that specifically binds atarget compound or moiety. The term “nucleic acid enzyme” (NAE) refersto nucleic acid that catalyses a chemical reaction (such as cleavage ofa substrate) when it binds a specific cofactor (such as a divalent metalion). Both an aptamer and a nucleic acid enzyme are examples ofreactors.

The term “conformational change” refers to the process by which anucleic acid, such as an aptamer, adopts a different secondary ortertiary structure. The term “fold” may be substituted forconformational change.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 illustrates an elevational view of a sensor base.

FIG. 2 illustrates a plan view of a sensor base.

FIG. 3 illustrates an opposing view to FIG. 2 of a sensor base.

FIG. 4 illustrates an elevational view of a sensor cap.

FIG. 5 illustrates a plan view of a sensor cap.

FIG. 6 illustrates an opposing view to FIG. 5 of a sensor cap.

FIG. 7 illustrates an elevational view of a cuvette.

FIG. 8 illustrates a syringe.

FIG. 9 illustrates a bag which contains a plurality of sensor housings.

DETAILED DESCRIPTION

The present invention makes use of the discovery of a compact reagentcontainer for field use. The compact reagent container preferablyincludes analysis chemistry reagents, such as nucleic acid enzymes,which allow the device to be adapted to many different analytes.Concentrations of ions such as lead, uranium, copper, mercury, cadmium,silver, etc., may be detected as low as a part per billion (ppb) level.In addition, a nozzle, designed to accelerate a sample fluid through thecompact reagent container, aids in mixing the sample fluid with theanalysis chemistry reagents. When used with a standard fluorometer, thecompact reagent container is capable of simply, rapidly, andinexpensively measuring very small concentrations of analytes.

FIG. 1 illustrates an elevational view of a sensor base 100 havingaspects of the present invention. The sensor base 100 includes an upperbase body 110, a lower base body 115, a wide channel 120, a narrowchannel 125, a frit 130, an insertive interlocking region 140, a nozzle150, an outer skirt 160, and an inner skirt 165. The frit 130 includesdried reagents 135. The insertive interlocking region 140 includes abase baffle 145.

The upper base body 110 is disposed on the lower base body 115. Theinsertive interlocking region 140 is disposed on the upper base body110. The base baffle 145 is disposed on the upper portion of theinsertive interlocking region 140. The wide channel 120 is disposed inand extends axially through the center of the insertive interlockingregion 140 and the upper base body 110. The narrow channel 125 isdisposed below the wide channel 120 and is in fluid connection with thewide channel 120. The narrow channel 125 extends axially through thecenter of the upper base body 110, the lower base body 115, and thenozzle 150. The frit 130 is disposed in the wide channel 120. The driedreagents 135 are disposed on the frit 130. The outer skirt 160 forms theperimeter of the lower base body 115 and extends along the height of thelower base body 115. The inner skirt 165 is disposed within the outerskirt 160 and surrounds the nozzle 150.

In one aspect, the dimensions of the sensor base 100 are 15 mm×15 mm×22mm. The lower base body 115 has a height of 14 mm. The upper base body110 has a height of 6 mm and a diameter of 12 mm. The insertiveinterlocking region 140 has a height of 2 mm and a diameter of 9 mm. Thebase baffle 145 has a height of 1 mm. The wide channel 120 has adiameter of 4.5 mm. The narrow channel 125 has a diameter of 2 mm.Preferably, the cross-sectional area of the wide channel 120 at the frit130 is greater than the cross-sectional area of the narrow channel 125at the nozzle 150.

The frit 130 preferably contains dried reagents 135 such as analysischemistry reagents. Such analysis chemistry reagents may include anucleic acid or nucleic acid enzyme. Examples of useful analysischemistry reagents can be found in U.S. Patent Application Publication,Pub. No. 2007/0269821, entitled “LATERAL FLOW DEVICES” to Mazumdar etal.; U.S. Pat. No. 6,706,474, entitled “NUCLEIC ACID ENZYME BIOSENSORSFOR IONS” to Lu et al.; U.S. Pat. No. 6,890,719, entitled “FLUORESCENCEBASED BIOSENSOR” to Lu et al.; International Publication Number WO2009/012309, entitled “NUCLEIC ACID BASED FLUORESCENT SENSOR FOR COPPERDETECTION” to Lu et al.; International Publication Number WO2009/045632, entitled “NUCLEIC ACID BASED FLUORESCENT SENSOR FOR MERCURYDETECTION” to Lu et al.; and U.S. Patent Application Publication, Pub.No. 2010/0151579, entitled “FLUORESCENT SENSOR FOR MERCURY” to Wang etal. The analysis chemistry reagents of these examples containpolynucleotides, such as nucleic acid enzymes, aptamers, aptazymes,and/or substrates; fluorophors; and quenchers. The visualization speciesand labels are fluorescent. U.S. Pat. No. 6,890,719 (noted above)describes analysis chemistry reagents including a nucleic acid enzymeand a substrate for the nucleic acid enzyme, each having a quencher,with one having a fluorophore. Other examples of useful analysischemistry reagents are described in U.S. Patent Application Publication,Pub. No. 2006/0094026, entitled “NUCLEIC ACID ENZYME LIGHT-UP SENSORUTILIZING INVASIVE DNA” to Lu et al.; and U.S. Patent ApplicationPublication, Pub. No. 2007/0037171, entitled “APTAMER-BASED COLORIMETRICSENSOR SYSTEMS” to Lu et al. These latter examples contain particles,and produce visualization species and labels that are colored.Alternatively, the visualization species and labels may be or a magnetic(such as a magnetic particle).

Preferably, the dried reagents 135 include one or more saccharides. Thesaccharide may be used to preserve the analysis chemistry reagents. Thesaccharide may be any water-soluble saccharide, includingmonosaccharides, disaccharides, and polysaccharides. Preferablemonosaccharides may include mannose, fructose, or ribose. Preferabledisaccharides may include trehalose, lactose, maltose, sucrose, orturanose. Preferable polysaccharides may include hydroxyethylstarch,inulin, or dextran. At present, a preferred saccharide is thedisaccharide trehalose. Preferably, the amount of saccharide present inthe dried reagent is 15 to 45% by weight, including 20, 25, 30, 35 and40% by weight. Other chemicals may be present in the dried reagents,such as a buffer or salts. The dried reagent should be protected fromcontact with skin, and should be kept dry before use. The dried reagentsmay be prepared by mixing one or more sugars, the analysis chemistryreagents, optionally a buffer and optionally salts, and then allowingthe mixture to air dry on the frit. Vacuum desiccation may then be usedto further dry the dried reagents.

The frit typically comprises a porous material, such as a porous plasticmedia (e.g., polyethylene), or cellulose, having pore sizes in the rangeof 10-50 microns, in order to remove any particulates in the sample. Theporous material of the frit may have a lower melting or degradationtemperature than sensor base.

FIG. 2 illustrates a plan view of a sensor base 100 having aspects ofthe present invention. The sensor base 100 includes an upper base body110, a lower base body 115, a wide channel 120, a frit 130, and aninsertive interlocking region 140. The insertive interlocking region 140includes a base baffle 145.

FIG. 3 illustrates an opposing view to FIG. 2 of a sensor base 100having aspects of the present invention. The sensor base 100 includes alower base body 115, a narrow channel 125, a nozzle 150, an outer skirt160, and an inner skirt 165.

FIG. 4 illustrates an elevational view of a sensor cap 400 havingaspects of the present invention. The sensor cap 400 includes an uppercap body 410, a lower cap body 415, a cap channel 420, a cap baffle 430,and a receptive interlocking region 440.

The upper cap body 410 is disposed on the lower cap body 415. The capbaffle 430 is disposed on the upper portion of the upper cap body 410.The cap channel 420 is disposed in and extends axially through thecenter of the upper cap body 410 and the lower cap body 415. Thediameter of the cap channel 420 decreases as it extends from the uppercap body 410 to the lower cap body 415. The receptive interlockingregion 440 is disposed in the lower portion of the lower cap body 415.

FIG. 5 illustrates a plan view of a sensor cap 400 having aspects of thepresent invention. The sensor cap 400 includes an upper cap body 410, alower cap body 415, a cap channel 420, and a cap baffle 430. The capbaffle 430 includes baffle flares 435. The baffle flares 435 aredisposed on opposing sides of the cap baffle 430.

FIG. 6 illustrates an opposing view to FIG. 5 of a sensor cap 400 havingaspects of the present invention. The sensor cap 400 includes a lowercap body 415, a cap channel 420, and a receptive interlocking region440.

In one aspect, the sensor cap 400 has a diameter of 12.5 mm and a heightof 18 mm. The diameter of the cap baffle 430 is 6.5 mm. At the capbaffle 430, the diameter of the cap channel 420 is 4 mm. At thereceptive interlocking region 440, the diameter of the cap channel 420is 3 mm. The height of the lower cap body 415 is 12 mm, while the heightof the upper cap body 410 is 6 mm. The diameter of the upper cap body410 is 5.5 mm.

The sensor base 100 and sensor cap 400 preferably comprise a durable andinert polymeric material. Preferred polymeric materials include, forexample, high-density polyethylene, low-density polyethylene,polypropylene, polycarbonate, polyethylene terephthalate, andpolystyrene. Fabrication of the sensor base 100 and sensor cap 400 maybe accomplished, for example, with injection molding.

FIG. 7 illustrates an elevational view of a cuvette 700 for use with thepresent invention. The cuvette 700 includes an upper chamber 710, alower chamber 715, cuvette opening 720, and orienting slots 730. Theupper chamber 710 is fluidly connected to the lower chamber 715. Thecuvette opening 720 is located at the upper portion of the upper chamber710. The orienting slots 730 extend vertically along front and rearfaces of the cuvette 700 and are located to the right and left of thelower chamber 715.

In operation, the cuvette 700 is inserted into a standard fluorometer.Preferably, the orienting slots 730 of the cuvette 700 fit a samplecompartment of the fluorometer and prevent the cuvette 700 from beinginserted into the fluorometer in an incorrect orientation.

In one aspect, the dimensions of the cuvette 700 are 45 mm×12 mm×12 mm.The width of the upper chamber 710 is 10 mm, while the width of thelower chamber 715 is 4 mm. The cuvette 700 preferably comprises atransparent material. Preferred materials include polycarbonate andglass. The cuvette 700 may be packaged and sold with the sensor base 100and/or sensor cap 400, or it may be packaged and sold by itself ineither individual or high quantity packaging.

FIG. 8 illustrates a syringe 800 for use with the present invention. Thesyringe 800 includes a barrel 810, a plunger 820, a rubber bulb 825, anda syringe tip 830. The rubber bulb 825 is coupled to the end of theplunger 820. The plunger 820 is slidably connected to the interior wallof the barrel 810 by the rubber bulb 825. The rubber bulb 825 forms aseal with the interior wall of the barrel 810. The syringe tip 830 is influid connection with the barrel 810.

In operation, the sensor cap 400 is placed on the sensor base 100. Theinsertive interlocking region 140 of the sensor base 100 is insertedinto the receptive interlocking region 440 of the sensor cap 400,mechanically coupling the sensor cap 400 to the sensor base 100 to forma sensor housing 910, illustrated in FIG. 9. The base baffle 145 fitssecurely into the receptive interlocking region 440 of the sensor cap,decreasing the incidence of fluid leakage between the sensor cap 400 andthe sensor base 100. The cuvette 700 is inserted into the lower basebody 115 of the sensor base 100. Upon insertion of the cuvette 700, theouter skirt 160 of the sensor base 100 surrounds the perimeter of theupper chamber 710 of the cuvette 700, while the inner skirt 165 and thenozzle 150 of the sensor base 100 are disposed within the cuvetteopening 720 of the upper chamber 710. When properly assembled, thesensor cap 400, sensor base 100, and cuvette 700 are in fluidconnection.

A fluid sample is first collected by the syringe 800, typically about 1ml. Depending upon the sample to be tested, additional sampleconditioning or preparation may be used, such as using an EPA protocolto obtain a sample of lead paint, or adding a buffering agent to adjustpH, and/or salts, which may be provided in a sample tube or calibrationtube. The syringe tip 830 is inserted into the cap channel 420 of thesensor cap 400. Preferably, the syringe tip 830 fits tightly into thecap channel 420 of the sensor cap 400 so as to decrease the incidence offluid leakage. Once the plunger 820 of the syringe 800 is depressed,fluid travels through the cap channel 420 and into the wide channel 120of the sensor base 100. As the fluid travels through the frit 130, itcontacts and re-hydrates the dried reagents 135. The fluid sample andthe re-hydrated reagents then flow into the narrow channel 125, out ofthe nozzle 150, through the upper chamber 710 of the cuvette 700, andinto the lower chamber 715 of the cuvette 700. In conjunction with thecap channel 420, wide channel 120, and narrow channel 125, the nozzle150 is designed to accelerate the sample fluid through the sensorhousing 910 and into the cuvette 700. Such acceleration aids in mixingthe sample fluid with the reagents. Once the reagents are mixed andreacting with the sample fluid in the upper chamber 710 and/or lowerchamber 715 of the cuvette 700, the cuvette 700 can be inserted into andanalyzed by a standard fluorometer. The cuvette 700 may be inserted intoa standard fluorometer prior to filling with fluids.

FIG. 9 illustrates a kit 900 having aspects of the present invention.The kit 900 includes a box 910, which contains a first bag 920 and asecond bag 922. The first bag 920 contains a plurality of black bags924, a plurality of sample tubes 940, and a plurality of syringes 800.Each black bag 924 in the first bag 920 contains a sensor housing 930, acuvette 700, and a desiccant (not shown in FIG. 9). Each sensor housing930 includes a sensor base 100 and a sensor cap 400. The second bag 922contains three black bags 924, three sample tubes 940, three calibrationtubes 950, and three syringes 800. Each black bag 924 in the second bag922 contains a calibration sensor 935, a cuvette 700, and a desiccant(not shown in FIG. 9). The calibration sensor 935 includes a sensor base100 and a sensor cap 400. Preferably, each black bag 924 blocks lightfrom impinging upon the sensor housings 930 and/or calibration sensors935.

The calibration sensor 935 may be provided with a calibration tube 950,which may containing a buffer and a known amount of the analyte ofinterest, for example lead with a buffer for pH 7, or UO₂ ²⁺ with abuffer for pH 5, and is used to calibrate a fluorometer. A sample tube940 may be provided with each sensor housing 930, and also may contain abuffer. Other agents that may also be provided in the calibration and/orsample tube include salts, and organic chelators. The reagents presentin the sample tube and/or calibration tube may be present in liquid formas a solution, or as a dry pellet or powders. The dried reagent on thefrit of the calibration sensor 935 may also include additional buffer tocompensate for acidity of the calibration amount of analyte. Examples ofbiological buffers include MES, HEPES, Tris, MOPS etc., and examples ofsalts include sodium chloride, potassium chloride, ammonium chloride,magnesium chloride, calcium chloride etc. A calibration standard couldalso be provided as a solution in a bottle, or in an ampoule.

The sensor housings 930 may be sold and/or distributed in bulk. Forexample, a plurality of sensor housings 930 may be distributed in bagswhich contain twenty-five, fifty, or one hundred sensor housings 930.Alternatively, the sensor bases 100 and sensor caps 400 of the sensorhousings 930 may be distributed separately. The sensor base and thesensor cap may be snapped together, or ultrasonically welded together,to help minimize any leakage.

An example of a kit is a box, containing sensor housings, calibrationsensor housing, syringes, sample tubes, calibration tubes, and cuvettes.The sensor housings may each be packaged in a bag, such as a black bagthat keeps out light, along with a cuvette, and a desiccant. Thecalibration sensor housings may each be packaged in a bag, such as ablack bag that keeps out light, along with a cuvette, and a desiccant.The bags of calibration sensor housing may be packed into a larger bag,which contain calibration tubes and syringes for use with thecalibration sensors. The bags of sensor housing may similarly be packedinto a larger bag, which contains sample tubes and syringes for use withthe sensors. The calibration and sample tubes may be 5 ml or 20 mltubes, for example.

The sensor housings 930 may be color-coded to signify the analyte whichthey are capable of detecting. In an exemplary embodiment, sensorhousings 930 are color-coded to correspond to selected analytes, such asgreen for lead, orange for uranium, blue for copper, etc., with darkeror lighter corresponding colors utilized for the sensor housings 930used in the corresponding calibration of the selected analyte.Preferably, the calibration sensor 935 for a particular analyte iscolored a different shade than the sensor housings 930.

Example

The following is an example of using a sensor housing:

Open the black bag which contains sensor in green plastic housing andplastic cuvette for lead analysis. The bag contains a transparentdesiccant pouch which should be discarded. Place the plastic cuvette inthe instrument, and then place the sensor housing on the cuvette.Collect test water in a clean cup, then slowly pour the test water intothe provided sample tube containing liquid buffer up to the 5 mL mark.Close the cap tightly and mix well by shaking. Next, draw 1 ml of waterfrom the sample tube into a syringe. Attach the syringe to the housingover the cuvette in the instrument. Squeeze water through housing intothe cuvette, then quickly remove the housing and syringe, close thesample chamber door of the instrument.

1. A sensor, comprising: a sensor housing, having a channel; a poroussubstrate, in the channel; an analysis chemistry reagent, on the poroussubstrate; and a nozzle, in fluid connection with the channel; whereinthe porous substrate fills a cross section of the channel, and thecross-sectional area of the channel at the porous substrate is greaterthan the cross-sectional area at the nozzle.
 2. The sensor of claim 1,wherein the analysis chemistry reagent comprises a nucleic acid enzyme.3. The sensor of claim 1, wherein the porous substrate is a frit.
 4. Thesensor of claim 1, wherein the sensor housing comprises: a sensor cap;and a sensor base; wherein the channel extends axially through thesensor cap and sensor base.
 5. The sensor of claim 4, furthercomprising: a first interlocking region, on the sensor cap; and a secondinterlocking region, on the sensor base; wherein the first interlockingregion couples to the second interlocking region to attach the sensorcap to the sensor base.
 6. The sensor of claim 1, wherein the analysischemistry reagent comprises a fluorophore.
 7. The sensor of claim 3,wherein the frit comprises cellulose or plastic.
 8. The sensor of claim4, wherein the sensor cap and the sensor base have been welded together.9. The sensor of claim 1, wherein the analysis chemistry reagentcomprises a saccharide.
 10. The sensor of claim 1, wherein the analysischemistry reagent comprises a nucleic acid enzyme, a fluorophore and asaccharide, the porous substrate is a frit comprising cellulose orplastic, and the sensor housing comprises: a sensor cap; and a sensorbase; wherein the channel extends axially through the sensor cap andsensor base.
 11. A kit, comprising: a box; and a plurality of thesensors of claim 1, and a plurality of cuvettes; wherein one of theplurality of sensors and one of the plurality of cuvettes are each in afirst sealed bag of a plurality of first sealed bags.
 12. The kit ofclaim 11, further comprising a plurality of syringes.
 13. The kit ofclaim 11, further comprising a plurality of sample tubes, each sampletube containing a buffer.
 14. The kit of claim 13, wherein the buffer ispresent as a dry solid.
 15. The kit of claim 11, further comprising aplurality of calibration sensors, wherein one of the plurality ofcalibration sensors and one of the plurality of cuvettes are each in asecond sealed bag of a plurality of second sealed bags.
 16. The kit ofclaim 15, further comprising a plurality of calibration tubes, eachcalibration tube containing a buffer and standard.
 17. The kit of claim16, further comprising a third sealed bag, wherein the plurality ofsecond sealed bags and the plurality of calibration tubes are in thethird sealed bag.
 18. The kit of claim 11, further comprising: aplurality of syringes, a plurality of sample tubes, each sample tubecontaining a buffer, a plurality of calibration sensors, wherein one ofthe plurality of calibration sensors and one of the plurality ofcuvettes are each in a second sealed bag of a plurality of second sealedbags, a plurality of calibration tubes, each calibration tube containinga buffer and standard, wherein the plurality of second sealed bags andthe plurality of calibration tubes are in the third sealed bag.
 19. Amethod of detecting an ion, comprising: flowing a sample fluid throughthe channel of the sensor of claim 1, to produce a product; collectingthe product; and detecting the presence of the product.
 20. The methodof claim 19, wherein the product is detected using fluorescencespectroscopy.